Project Summary The steady increase of human lifespan has led to a dramatic increase in the elderly population. There is unequivocal evidence from humans and animal models that the aging process predisposes to hepatic functional impairment, resulting in significantly increased susceptibility to acute and chronic liver injury. For an example, acetaminophen remains the first-line drug widely used to manage mild to moderate pain in the elderly population. Accidental acetaminophen hepatotoxicity, the most common drug-induced liver injury (DILI), accounts for ~15% of all DILI in the general population but ~55% of all DILI in the elderly population. Furthermore, accidental acetaminophen hepatotoxicity in the elderly generally shows poor outcome. Currently, aging-associated liver pathophysiology contributing to hypersensitivity and poor prognosis of drug-induced hepatotoxicity is still not well understood. Bile acid signaling play important roles in regulating liver metabolic homeostasis. Hepatic bile acid signaling is known to be significantly impaired in aging, but its implication in aging-associated liver impairment is still not clear. The goal of this study is to investigate a novel mechanism by which impaired bile acid signaling disrupted hepatic cysteine, glutathione (GSH) and coenzyme A (CoA) homeostasis, which renders the liver more susceptible to external insults in aging. The amino acid cysteine supports hepatic synthesis of GSH and CoA. GSH plays a critical role in detoxification of drugs and environmental toxins, and its hepatic and plasma concentration decreases significantly with age. CoA is a key cofactor required to support hepatic catabolic metabolism. CoA sequestration and depletion by drugs and their metabolites has been linked to mitochondrial impairment and liver injury. This project is developed based on our discovery that under normal physiology bile acids exert a strong repression on the cysteine dioxygenase type-1 (CDO1) - mediated cysteine elimination pathway, which is important in maintaining hepatic free cysteine pool. Impaired bile acid signaling such as in aging results in markedly increased liver CDO1 expression and a hepatic cysteine, GSH and CoA deficient state, which renders the liver more vulnerable to oxidative and metabolic stress-induced injury. Three specific aims are proposed to investigate the significance of the bile acid-CDO1-csyteine axis in modulating acetaminophen-induced liver injury in young adult and aged mice. This study will employ state-of-the-art models and techniques including AAV8-mediated CDO1 gene delivery, targeted metabolic profiling, tracer-mediated metabolic flux, and Seahorse Flux Analyzer measurement of mitochondrial function to interrogate how CDO1-mediated cysteine metabolic flux modulates hepatic redox homeostasis, organelle function and sensitivity to injury.